| Přispěvatelé: |
Trajković, Vladimir, Harhaji-Trajković, Ljubica, Isaković, Aleksandra, Andrić, Silvana, Tovilović Kovačević, Gordana |
| Popis: |
Autofagija, razgradnja nepotrebnih/nefunkcionalnih unutarćelijskih komponenti u autolizozomima, kiselim organelama nastalim spajanjem autofagozoma i lizozoma, može biti citoprotektivna i citotoksična. U ovoj tezi ispitivana je uloga glavnog ćelijskog energetskog senzora protein kinaze aktivirane adenozin monofosfatom (AMPK) i mitogenom aktivirane protein (MAP) kinaze p38 u autofagiji i apoptozi ćelija humanog neuroblastoma SH-SY5Y izazvanoj mimetikom Parkinsonove bolesti 6-hidroksidopaminom (6-OHDA). 6-OHDA je indukovao apoptozu zavisnu od oksidativnog stresa i aktivacije kaspaza. Prisustvo autofagnih vezikula, zakišeljavanje citoplazme, autofagozomima asocirana konverzija LC3 proteina i razgradnja supstrata autofagne proteolize p62 ukazali su da 6-OHDA indukuje autofagiju. 6-OHDA je aktivirao AMPK i njegov supstrat Raptor, te inhibirao glavni supresor autofagije mTOR i njegov supstrat S6K, uprkos tome što je stimulisao mTOR aktivator Akt. Konverzija LC3, degradacija p62, zakišeljavanje citoplazme i inhibicija mTOR/S6K izazvani 6-OHDA poništeni su supresijom AMPK. Inhibicija AMPK i autofagije su smanjile, dok su inhibicije mTOR i Akt pojačale oksidativni stres i apoptozu indukovanu 6-OHDA. 6-OHDA je stimulisao MAP kinaze JNK, ERK i p38. Inhibicija JNK i ERK nisu imale uticaja, dok je inhibicija p38 suprimirala proapoptotsko dejstvo 6-OHDA, ali nije delovala na aktivnost AMPK i autofagiju. Sa druge strane, inhibicija AMPK smanjila je aktivaciju p38 u ćelijama tretiranim 6-OHDA. Antioksidans N-acetil cistein je inhibirao aktivaciju AMPK, p38 i autofagiju stimulisanu 6-OHDA. Navedeni rezultati ukazuju da bi oksidativnim stresom indukovana AMPK/mTOR zavisna citotoksična autofagija i AMPK/p38 zavisna apoptoza mogle biti pogodne mete za terapiju Parkinsonove bolesti. Autophagy, the degradation of unused/dysfunctional cellular components in autolysosomes, acidic organelles created by fusion of autophagosomes and lysosomes, can be either cytotoxic or cytoprotective. Here we investigated the role of the main cellular energy sensor, AMP-activated protein kinase (AMPK) and mitogen activated protein (MAP) kinase p38 in autophagy and apoptosis caused by the Parkinsonian mimetic 6-hydroxydopamine (6-OHDA) in human neuroblastoma SH-SY5Y cells. 6-OHDA induced apoptosis dependent on oxidative stress and caspase activation. Presence of autophagic vesicles, cytoplasm acidification, autophagosomeassociated conversion of LC3 protein, degradation of autophagic proteolysis substrate p62, all indicated that 6-OHDA induced autohagy. 6-OHDA activated AMPK and its substrate Raptor, and suppressed main autophagy inhibitor, mTOR and its target S6K, in spite of activating the mTORactivating Akt. LC3 conversion, p62 degradation, cytoplasm acidification and mTOR/S6K inhibition caused by 6-OHDA were all abolished by AMPK suppression. Inhibition of AMPK and autophagy decreased, while inhibition of mTOR and Akt potentiated oxidative stress and apoptosis caused by 6-OHDA. 6-OHDA stimulated MAP kinases JNK, ERK and p38. Inhibition of JNK and ERK did not affect, while p38 inhibition reduced pro-apoptotic effects of 6-OHDA, although it did not influence AMPK activation nor autophagy. Conversely, AMPK inhibition mitigated p38 activation in 6-OHDA treated cells. Antioxidant N-acetyl cysteine suppressed activation of AMPK, p38 and autophagy stimulated by 6-OHDA. These results suggest that oxidative stress-induced, AMPK/mTOR-dependent cytotoxic autophagy and AMPK/p38-dependent apoptosis could be valid therapeutic targets for treating Parkinson’s disease. |
